Florence Auguy

Rice yellow mottle virus (RYMV) is one of the most devastating viral pathogens affecting rice cultivation in Africa. The eukaryotic translation initiation factor 4G isoform eIFiso4G1 plays a pivotal role in rice susceptibility to the virus. Naturally occurring resistance alleles impair infection but are predominantly found in the African cultivated species Oryza glaberrima , whose use in breeding programs is limited by interspecific sterility barriers with the widely grown O. sativa . Here, we used CRISPR/Cas9 to generate knockout (KO) mutations in eIFiso4G1 as well as insertion-deletion (Indel) variants within the region involved in interaction with the viral protein VPg. KO mutations conferred high resistance to RYMV. No resistance-breaking events were observed, suggesting that this resistance may be more durable than that conferred by naturally occurring alleles. However, complete gene KO slightly affected plant growth. Lines carrying Indel mutations in the VPg-interacting region displayed variable resistance levels, with some variants conferring high resistance without compromising plant development. Structural modeling of the eIFiso4G1 variants and their complexes with VPg provided mechanistic insights into how specific Indels modulate the resistance phenotype.

Xanthomonas oryzae pv. oryzae ( Xoo) causes bacterial leaf blight (BLB), a major rice disease causing up to 70% yield loss in Asia and West Africa. First described in Japan in 1884 and later reported in West Africa in the 1970s, BLB recently emerged in East Africa, with an epidemic reported in Tanzania in 2019. Remarkably, the disease was detected for the first time in Madagascar the same year, representing a serious threat to food security. To investigate the origin of BLB in Madagascar, we isolated 73 Xoo strains from symptomatic rice leaves collected between 2019 and 2023. Multilocus variable-number of tandem-repeats analysis genotyping revealed 19 haplotypes forming a single clonal complex, indicating low diversity and a likely recent introduction. To come up with disease control strategies, IRBB-based race-typing was achieved and identified four resistance genes ( Xa8, xa13, Xa21, and Xa23) that confer resistance to all Malagasy strains tested, whereas the 19 Malagasy varieties assessed were susceptible. The analysis of SWEET knockout lines confirmed that Malagasy strains rely on the susceptibility gene OsSWEET11 for full virulence. Whole-genome sequencing and transcription activator-like effector repertoire analyses of two strains allowed for the identification of a PthXo1 ortholog predicted to induce OsSWEET11. Single-nucleotide polymorphism-based phylogenetic analyses clustered Malagasy strains within Asian lineages, most closely related to strains originated from India. Malagasy strains did not cluster with recently reported Tanzanian Xoo, suggesting independent introductions. Overall, our study demonstrates that BLB in Madagascar results from a recent and single introduction from Asia and identifies effective resistance genes for deployment. [ ] Copyright © 2026 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Rice yellow mottle virus (RYMV) is one of the most devastating viral pathogens affecting rice cultivation in Africa. The eukaryotic translation initiation factor 4G isoform eIFiso4G1 plays a pivotal role in rice susceptibility to the virus. Naturally occuring resistance alleles impair infection but are predominantly found in the African cultivated species Oryza glaberrima, whose use in breeding programs is limited by interspecific sterility barriers with the widely grown O. sativa. Here, we used CRISPR-Cas9 to generate knockout (KO) mutations in eIFiso4G1 as well as insertion-deletion (Indel) variants within the region involved in interaction with the viral protein VPg. KO mutations conferred high resistance to RYMV. No resistance-breaking events were observed, suggesting that this resistance may be more durable than that conferred by naturally occuring alleles. However, complete gene KO slightly affected plant growth. Lines carrying Indel mutations in the VPg-interacting region displayed variable resistance levels, with some variants conferring high resistance without compromising plant development. Structural modeling of the eIFiso4G1 variants and their complexes with VPg provided mechanistic insights into how specific Indels modulate the resistance phenotype.

Bacterial leaf blight (BLB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is a devastating disease of rice crops which can lead to yield losses ranging from 20 to 30% and as high as 50% (Mew 1987). BLB was first reported in Japan in 1884 and is now prevalent in most of the rice-producing countries in Asia. In Africa, Xoo was first reported in Mali and Cameroon (Buddenhagen et al., 1979) and later in many West African countries. Recently, BLB also appears to be spreading in East Africa with reports in Tanzania (Schepler-Luu et al., 2023) and in Madagascar (Raveloson et al., 2023). In June 2023, typical symptoms of BLB such as water-soaked areas and yellowish lesions along the leaf margins and tips were observed on rice plants in the coastal region of Kenya. Five rice fields of the Taita Taveta County with an incidence of 5-10% were surveyed, and 10-15 symptomatic leaf samples were collected in each field following a W pattern. Rice varieties were either Saro 5 or Arize 6444, two high productivity rice. 4-5cm long leaf fragments of 45 samples were next processed for bacterial isolation and surface disinfected, rinsed with sterile water, and ground individually in tubes with steel beads using a TissueLyser (Qiagen). Thirty-four round, pale yellow pigmented, and mucoid Xanthomonas-like colonies were obtained upon plating of 30 µl of the leaf powder resuspended in 1mL of sterile water on a semi-selective medium (peptone, 10 g/L; sucrose, 10 g/L; purified bacteriological agar, 16 g/L; glutamic acid, 1g/L; cycloheximide, 25 mg/L; cephalexin, 20 mg/L and kasugamycin, 10 mg/L) 72h after incubation at 28°C. To confirm that the isolates were Xoo, multiplex PCR established for the identification of X. oryzae pathovars was used (Lang et al. 2010). Thirty-two strains resulted in the amplification of two bands characteristic of the Xoo pattern. In addition, 6 of these strains (namely CIX5498, CIX5504, CIX5513, CIX5514, CIX5516 and CIX5539) were subjected to gyrB sequencing using the universal primers XgyrB1F and XgyrB1R (Young et al., 2008). The resulting partial 800 bp gyrB sequences of these 6 strains showed 100% identity (767/767 nt) with that of the PX099A Asian reference Xoo strain (accession CP000967.2). The pathogenicity of these 6 strains and the reference strain PX099A used as positive control was tested on 1-month-old Oryza sativa cv. Komboka plants (n=8 per treatment, 1 repetition). Rice leaves were inoculated by leaf clipping using sterile scissors dipped in the bacterial suspension at an optical density of 0.2. Three weeks after inoculation, plants exhibited typical BLB lesions for each of the six tested strains similarly to that of the positive control, whereas Mock-inoculated plants remained symptomless. Bacteria re-isolated from these diseased leaves yielded colonies that were confirmed as Xoo by multiplex PCR, fulfilling Koch's postulate. All strains isolated in this study are referenced in the CIX collection of the French National Research Institute for Sustainable Development in Montpellier. To our knowledge, this study represents the first molecular confirmation of the presence of BLB in Kenya and complement our knowledge of this important disease in East Africa. This work will serve as a basis for future rice breeding for BLB resistance efforts in Kenya. Further surveys are needed to clarify the spatial distribution and prevalence of BLB in other regions of Kenya and further analysis will be needed to understand the origin of the outbreak and to identify strategies to control BLB in Kenya.

Summary: Rice yellow mottle virus (RYMV) causes one of the most devastating rice diseases in Africa. Management of RYMV is challenging. Genetic resistance provides the most effective and environment-friendly control. The recessive resistance locus rymv2 ( OsCPR5 . 1 ) had been identified in African rice ( O. glaberrima ), however, introgression into O. sativa ssp. japonica and indica remains challenging due to crossing barriers. Here, we evaluated whether CRISPR/Cas9 genome editing of the two rice nucleoporin paralogs OsCPR5 . 1 ( RYMV2 ) and OsCPR5 . 2 can be used to introduce RYMV resistance into the japonica variety Kitaake. Both paralogs had been shown to complement the defects of the Arabidopsis atcpr5 mutant, indicating partial redundancy. Despite striking sequence and structural similarities between the two paralogs, only o scpr5 . 1 loss-of-function mutants were fully resistant, while loss-of-function oscpr5 . 2 mutants remained susceptible, intimating that OsCPR5 . 1 plays a specific role in RYMV susceptibility. Notably, edited lines with short in-frame deletions or replacements in the N-terminal domain (predicted to be unstructured) of OsCPR5 . 1 were hypersusceptible to RYMV. In contrast to mutations in the single Arabidopsis AtCPR5 gene, which caused severely dwarfed plants, oscpr5 . 1 and oscpr5 . 2 single knockout mutants show neither substantial growth defects nor symptoms indicative of programmed cell death, possibly reflecting functional redundancy of the isoforms regarding other important functions. The specific editing of OsCPR5 . 1 , while maintaining OsCPR5 . 2 activity, provides a promising strategy for generating RYMV-resistance in elite Oryza sativa lines as well as for effective stacking with other RYMV resistance genes or other traits.

Xanthomonas oryzae pv. oryzae (Xoo) strains that cause bacterial leaf blight (BLB) limit rice (Oryza sativa) production and require breeding more resistant varieties. Transcription activator-like effectors (TALEs) activate transcription to promote leaf colonization by binding to specific plant host DNA sequences termed effector binding elements (EBEs). Xoo major TALEs universally target susceptibility genes of the SWEET transporter family. TALE-unresponsive alleles of clade III OsSWEET susceptibility gene promoter created with genome editing confer broad resistance on Asian Xoo strains. African Xoo strains rely primarily on the major TALE TalC, which targets OsSWEET14. Although the virulence of a talC mutant strain is severely impaired, abrogating OsSWEET14 induction with genome editing does not confer equivalent resistance on African Xoo. To address this contradiction, we postulated the existence of a TalC target susceptibility gene redundant with OsSWEET14. Bioinformatics analysis identified a rice locus named ATAC composed of the INCREASED LEAF INCLINATION 2 (ILI2) gene and a putative lncRNA that are shown to be bidirectionally upregulated in a TalC-dependent fashion. Gain-of-function approaches with designer TALEs inducing ATAC sequences did not complement the virulence of a Xoo strain defective for SWEET gene activation. While editing the TalC EBE at the ATAC loci compromised TalC-mediated induction, multiplex edited lines with mutations at the OsSWEET14 and ATAC loci remained essentially susceptible to African Xoo strains. Overall, this work indicates that ATAC is a probable TalC off-target locus but nonetheless documents the first example of divergent transcription activation by a native TALE during infection.

Non-coding small RNAs (sRNA) act as mediators of gene silencing and regulate plant growth, development and stress responses. Early insights into plant sRNAs established a role in antiviral defense and they are now extensively studied across plant-microbe interactions. Here, sRNA sequencing discovered a class of sRNA in rice ( Oryza sativa ) specifically associated with foliar diseases caused by Xanthomonas oryzae bacteria. Xanthomonas -induced small RNAs (xisRNAs) loci were distinctively upregulated in response to diverse virulent strains at an early stage of infection producing a single duplex of 20-22nt sRNAs. xisRNAs production was dependent on the Type III secretion system, a major bacterial virulence factor for host colonization. xisRNA loci overlap with annotated transcripts sequences often encoding protein kinase domain proteins. A number of the corresponding rice cis- genes have documented functions in immune signaling and some xisRNA loci coincide with the coding sequence of a conserved kinase motif. xisRNAs exhibit features of small interfering RNAs and their biosynthesis depend on canonical components OsDCL1 and OsHEN1. xisRNA induction possibly mediates post-transcriptional gene silencing but they do not broadly suppress cis -genes expression on the basis of mRNA-seq data. Overall, our results identify a group of unusual sRNAs with a potential role in plant-microbe interactions.